The present invention relates to a polarographic cell provided with an immobilized enzyme membrane. More particularly, it relates to an enzyme electrode for detecting hydrogen peroxide comprising an anode, a cathode, an immobilized enzyme membrane having a specific structure and an electrolyte, said immobilized enzyme membrane being arranged facing to the anode.
There are widely used methods for determining selectively a slight amount of various components such as glucose, urea, uric acid, triglycerides, phospholipids, creatinine, amino acids, lactic acid, xanthine, chondroitin, transaminase, etc. which are contained in body fluid, body tissue, foodstuffs, or the like.
Generally, enzymes have a specificity against a substrate thereof and act selectively thereon under mild conditions, and hence, they are advantageously employed for the determination of such components as mentioned above without using any specific reagent and also without any problem of environmental pollution. The enzymes have, on ther other hand, some drawbacks that they are chemically and physically unstable and the process of the determination is so complicated. In order to eliminate the drawbacks, it has been proposed to immobilize the enzyme. Besides, it has also been proposed to employ the immobilized enzymes in a polarography which is useful for the electrical determination of components contained in a slight amount in test samples (cf. Japanese Patent Publication No. 28672/1972). That is, an enzyme electrode provided with an immobilized enzyme membrane is contacted with a solution to be analyzed, by which the enzyme is reacted with the substrate to produce an electrode detectable substance, e.g. hydrogen peroxide, and the resulting detectable substance is detected with an electrode cell. In this manner, the component contained in the test solution is polarographically analyzed. When the electrode detectable substance is hydrogen peroxide, the hydrogen peroxide is decomposed with the anode, whereby an electric current flows through the polarographic cell in proportion to the amount of hydrogen peroxide. Since the value of electric current is proportionate to the amount of the component to be determined, the amount of the component can be calculated. In practice of this method, however, it is necessary to previously remove the interfering materials contained in the test solution, which are active to the polarography. For instance, when glucose contained in blood is determined, other components active to the polarography, such as uric acid, ascorbic acid, glutathione, mercaptoacetic acid, etc., should previously be removed. In order to correct the error due to these interfering materials, it has been proposed to employ a multiple electrode system, whereby the signal owing to the interfering materials is corrected (cf. Japanese Patent Publication No. 35360/1970). This system has, however, complicated electrode constituents and electrical circuit, and hence, this system is disadvantageously expensive and troublesome in the operation thereof. It has also been proposed to employ a laminate membrane comprising a membrane which can selectively permeate the detectable substance, an immobilized enzyme membrane and a membrane for removing high molecular materials in order to eliminate low molecular weight interfering materials of (cf. Japanese Patent Laid Open Publication (unexamined) No. 55691/1977). However, it is troublesome to laminate and adhere multiple membranes uniformly and precisely, which requires a highly trained technique. Moreover, the laminate membrane has a low strength and is easily broken and shrunk, and hence, it is difficult to install onto the electrode or to take off therefrom. It has also been proposed to employ a porous membrane (cf. Japanese Patent Laid Open Publication (unexamined) No. 17889/1977). However, when the porous membrane has a small pore diameter, it shows a good selective permeability of the detectable substance, but the contact of the substrate and the enzyme becomes insufficient, which induces inaccurate results of the determination. On the other hand, when the porous membrane has a large pore diameter, the selective permeability becomes insufficient and the interfering materials also permeate, which induces inaccurate results of the determination, too.